Integrins are transmembrane receptors that bind extracellular matrix proteins or other adhesion receptors on neighboring cells. Heterodimeric pairing of integrin α and β subunits confers specificity of binding to one or more substrates (Weis et al., 2011, Cold Spring Harb Perspect Med; 1:a006478). This family of adhesion molecules plays a pivotal role in broad contexts of biology, including inflammation, innate and antigen specific immunity, homeostasis, wound healing, tissue morphogenesis, and regulation of cell growth and differentiation. Dysregulation of integrins is involved in the pathogenesis of many disease states, from autoimmunity to thrombotic vascular diseases to cancer metastasis. Extensive efforts have been directed towards the discovery and development of integrin antagonists for clinical applications.
The αv integrins, each having an αv subunit paired with a β1, β3, β5, β6 or (38 subunit, appear to be particularly important during the tissue remodeling associated with wound repair, angiogenesis, and cancer (Weis et al., 2011, supra). The αv integrins are being targeted for cancer, ophthalmological and orthopedic indications. Integrins αvβ3 and αvβ5 have also been associated with tumors, arthritis, psoriasis and age-related macular degeneration (AMD). In particular, αvβ3 integrin is important in mediating angiogenesis and in inhibiting tumor migration, and αvβ6 integrin is upregulated in some cancers. The other αv integrins present in the cornea (αvβ5, αvβ6, and αvβ8) mediate transforming growth factor β (TGFβ) activation.
It was reported that the integrins α5β1, αvβ3 and αvβ5 play an important role in the process of angiogenesis and are expressed in a variety of malignancies, including, but not limited to, melanoma, breast cancer, prostate cancer, colon cancer, and gliomas (Staunton et al., 2006, Adv Immunol., 91:111-57). The intratumoral expression of these integrins has been associated with progression and metastasis in tumors, such as melanoma, breast cancer, and prostate cancer (Staunton et al., 2006, supra). They have been shown to signal through multiple pathways and contribute to endothelial cell migration and proliferation. In vivo, they are overexpressed on tumor neovasculature and on tumor cells themselves, which suggests that their function may potentiate tumor progression by multiple mechanisms. Antagonistic antibodies and small molecules directed against integrins α5β1, αvβ3, and αvβ5 have been shown to inhibit angiogenesis in vitro and in vivo. Inhibitors of integrins α5β1, αvβ3, and αvβ5 are able to inhibit signaling through ERK, Akt and FAK, resulting in decreased adhesion, migration and proliferation of endothelial and cancer cells. These antagonists have also been found to elicit cell death through caspase-dependent mechanisms. Therefore, the critical role of integrinsα5β1, αvβ3 and αvβ5 in angiogenesis and association with tumor progression make them attractive targets for anticancer therapy, and many antagonists of these integrins have been tested in clinical trials.
The αvβ3 integrin shares the same β3 subunit with the αIIbβ3 integrin, as well as several macromolecular ligands including fibrinogen, fibronectin, thrombospondin, von Willebrand factor, and vitronectin. These ligands all contain a triple amino acid sequence arginine-glycine-aspartic acid (RGD). Fibronectin and vitronectin are also ligands for α5β1 and other αv integrins. The αIIbβ3 integrin is a major membrane protein on platelets and plays an important role in platelet aggregation. Several αIIbβ3 integrin antagonists have been developed for the treatment of patients with acute coronary syndrome (ACS). However, because extensive inhibition of platelet aggregation are associated with increased risk of bleeding, ongoing studies are focused on reduction of bleeding and other side effects of αIIbβ3 integrin antagonists. It is essential to design drugs by blocking either a single integrin or multiple αv integrins for different indications (Goodman, 2012. Trends Pharmacol Sci. 2012; 33:405-412).
Disintegrins are a family of low-molecular-weight RGD-containing peptides that bind to integrins, such as, αIIbβ3, α5β1, and αvβ3 expressed on platelets and other cells, including vascular endothelial cells and some tumor cells. In addition to their potent anti-platelet activity, studies of disintegrins have revealed new uses in the diagnosis of cardiovascular diseases and the design of therapeutic agents in arterial thrombosis, osteoporosis, and angiogenesis-related tumor growth and metastasis. Rhodostomin (Rho), a disintegrin from the venom of Colloselasma rhodostoma, has been found to inhibit platelet aggregation in vitro and in vivo through the blockade of platelet glycoprotein αIIbβ3. It is also found that Rho can bind to integrins αIIbβ3, α5β1, and αvβ3 with high affinity and interact with cancer cells. For example, Rho is reported to inhibit the adhesion of breast and prostate carcinoma cells to both unmineralized and mineralized bone extracellular matrices in a dose-dependent manner, without affecting the viability of tumor cells. Rho also inhibits the migration and invasion of breast and prostate carcinoma cells.
However, because rhodostomin non-specifically binds to integrins αIIbβ3, α5β1, and αvβ3, the pharmaceutical uses of rhodostomin may cause serious side effects, such as bleeding resulting from the inhibition of platelet aggregation. Therefore, a need exists in the art for a disintegrin variant that is selective for integrins α5β1 and αvβ3, but with reduced binding activity to αIIbβ3. Such a need is met by this invention.